WO1998024301A1

WO1998024301A1 - Seedless tomato and method for producing a seedless tomato, hybrid tomato plants capable of producing said seedless tomatoes and cultivation material therefore, and food products obtained from said seedless tomatoes

Info

Publication number: WO1998024301A1
Application number: PCT/NL1997/000668
Authority: WO
Inventors: Gerardus Johannes Aloijsius Van Vliet
Original assignee: WESTERN SEED ESPANA SA
Current assignee: WESTERN SEED ESPANA SA
Priority date: 1996-12-06
Filing date: 1997-12-05
Publication date: 1998-06-11
Anticipated expiration: 1999-06-06
Also published as: AU5417598A; NL1004722C2; US20020010953A1

Abstract

The present application relates to a method for obtaining a seedless tomato, a tomato plant carrying said seedless tomatoes or capable of carrying said seedless tomatoes, or cultivation material such as seed for such a tomato plant, comprising : (a) providing a first tomato plant, that is homozygote recessive with regard to the properties parthenocarpy (pk,pk) and functional sterility (fs,fs); (b) providing a second tomato plant, that is homozygote recessive with regard to the properties parthenocarpy (pk,pk) and functional sterility (fs,fs); (c) crossing the first and second tomato plant for the production of cultivation material, such as seed, which is homozygote recessive with regard to the properties parthenocarpy (pk,pk) and functional sterility (fs,fs); (d) optionally, the cultivation of the cultivation material thus obtained into a tomato plant capable of carrying the seedless tomatoes ; (e) optionally, cultivating said tomato plant until it carries the seedless tomatoes, and harvesting the seedless tomatoes thus obtained. Step (c) is carried out by scissoring or cutting, followed by scraping of the pollen tube closed by the phenotype of functional sterility. The obtained tomato plant is preferably a hybrid. The invention further relates to cultivation material for tomatoes such as seed, tomato plants as well as a seedless tomato, obtainable according to the abovementioned method, as well as a processed product on a tomato basis obtained from such seedless tomatoes. Finally, the invention relates to the parent lines as well as a method for obtaining these parent lines.

Description

Seedless tomato and method for producing a seedless tomato, hybrid tomato plants capable of producing said seedless tomatoes and cultivation material therefore, and food products obtained from said seedless tomatoes.

The present application relates to a seedless tomato, a method for obtaining said seedless tomato, as well as a method for processing the seedless tomato thus obtained to products, in particular food products.

In modern agriculture, in producing tomatoes, hybrid seeds and tomato plants are used which have highly specific, desired properties. These hybrids are obtained by commonly known cultivation techniques and plant breeding techniques, which make it possible to "design" the hybrids, i.e. to confer to the hybrid plants predetermined, desired properties, by choosing the (properties of) parent lines which by crossing (can) provide the hybrid seed. In known methods for obtaining hybrid seeds, usually inbred lines are used, obtained by self-fertilization (self-pollination) over several generations. This makes these parent lines genetically homozygote and identical, providing genetically uniform -albeit heterozygote- hybrid line combinations, which (can) combine the properties of the parent lines. In doing so, the aim is on the one hand to cross certain properties from the parent lines as purely as possible into the seed, on the other hand use is made of the known effect of heterosis or inbred growth, which provides properties regarding -inter alia- the growth of plants and fruits and thereby of the yield. This heterosis effect is obtained when/because the parent lines used are not related with respect to certain genetic properties ("lie far apart"). For a further description of plant breeding techniques in general, and tomatoes in particular, using classical cultivation techniques, including the formation of hybrids, reference is made to the known handbooks, the contents of which are incorporated herein by reference.

The tomatoes that are currently on the market have the disadvantage that they contain seeds (pips). The presence of these seeds for many consumers detracts from the attractiveness of the tomato. Also, in the preparation of a large number of products on the basis of tomatoes, in particular food products on the basis of tomatoes, the seeds have to be removed, for instance by sieving, optionally after prior pureeing, boiling or mashing, which involves further processing steps. This is true for both the preparation of products on industrial scale, such as puree, soups, juices or sauces on tomato basis, as for the household preparation of dishes or food products.

The object of the invention is to solve the abovementioned problems. The tomato flower consists of an ovary, above which there is a pistil. Around the pistil there are several stamen, that produce pollen. In the ovary, there are several pre-embryo's/embryo's which develop (after pollination with the pollen) into seeds.

The tomato plant can be considered as an "obligatory self-pollinator", which means that almost exclusively only its own pollen ends up on the stamen of the flowers of the same plant and thereby pollinates the pre-embryo's. As soon as pollinated pre-embryo's are formed in the ovary, the ovary starts to grow into a tomato (fruit) containing within it, at the same time, the developing seed. In principle no fruit will be formed when no seeds are developing in the ovary (for instance as a result of not-pollinated pre-embryo's).

In the tomato, a gene is known, called the PK gene, which codes for the property of parthenocarpy. This gene, when present as a double recessive (pk,pk) leads to developments of fruit (fruit flesh) without concomitant development of seed. [In the present description, as usual in genetics, the use of capital letters indicates a dominant (form of a) gene and the use of small letters denotes a recessive gene: PK,PK therefore denotes a homozygote dominant genotype; pk,PK and PK,pk denote heterozygote genotypes; and pk,pk denotes a homozygote recessive genotype. As commonly known, only the homozygote recessive genotype will generally provide the corresponding recessive phenotype.]

In nature, or in the greenhouse, (the phenotype of) parthenocarpy will only be partial; the factors which are responsible for the absence or partial presence of seeds are present on alleles. In partial parthenocarpy, seeds are formed in (another) part of the fruit, as the result of which the fruit will grow irregularly, which is undesirable. Partial parthenocarpy therefore leads to irregular forms of the fruit, the fruits are deformed.

When a fruit is formed on the basis of total parthenocarpy, no seed is developed within the fruit, with the result that the genes coding for parthenocarpy are not passed over to the next generation, so that the generational line is ended.

Because of this, the gene is extremely rare in nature. Also, (the phenotype of) total parthenocarpy leads to fruits without seed, which makes the production of seed impossible.

The property of parthenocarpy resides in several alleles. The phenotype of total parthenocarpy can only occur when the "mother" (i.e. the tomato plant, the flower of which is pollinated with pollen) as well as the "father" (i.e. the tomato plant that provides the pollen) are both double recessive with regard to the PK gene. This is because, in the fruit "in statu nascendi", there are several developing seeds, at least one of which may not become homozygote recessive if one of the parents is a heterozygote.

Partial parthenocarpy (in the developing tomato, as the result of at least one seed which is not double recessive with regard to the PK gene) makes the fruit develop in a deformed manner ("detracts from the form of the tomato").

Besides the PK gene in tomato, a gene is known which codes for the property of functional sterility (FS). A double recessive plant with regard to FS

(fs,fs) leads to a tomato plant with a pollen tube that is totally closed, so that the complete and fertile pollen cannot leave the pollen tube, not even by vibrating or other mechanical influences (bumble bees, insects or a vibrator).

The pollen of a tomato plant which has the double recessive (fs,fs) phenotype can only be released by physically opening the pollen tube by hand (by cutting or scissoring), after which - in practice - the pollen has to be removed by hand from the opened pollen tube, i.e. by scraping.

For fertilization of the same or another tomato plant, the pollen then has to be applied to the pistil of the flower, which in practice also must be carried out manually. In any other "natural" way (i.e. without the abovementioned human intervention) the pollen of a functional sterile flower is not released and therefore not available for fertilization of a pre-embryo. A double recessive, functionally sterile plant (fs,fs) therefore does not fertilize pre-embryo's, which ends the generational line so that the recessive genes for functional sterility are not passed on to the next generation. In nature, with a double recessive phenotype for functional sterility (fs,fs), no fertilization of the pre-embryo's will take place so that no fruit (tomato) will be formed.

It has now been found that tomatoes without seeds/pips can be produced with advantage using -very preferably hybrid- tomato plants which combine the recessive phenotype of parthenocaφy (pk,pk) with the recessive phenotype of functional sterility (fs,fs).

In nature, the genotype combination of double recessive parthenocaφy and double recessive functional sterility has no chance of survival (as no seed is formed), so that the genes are not passed onto the next generation.

By combining the genetic properties of functional sterility (on the basis of the double recessive gene fs) and parthenocaφy (on the basis of the double recessive gene pk) a totally seedless fruit is formed, which however can grow because of the property of parthenocaφy, forming an attractive, regularly formed, seed- less fruit. Such regularly fo:τned, absolutely seedless fruits cannot spontaneously occur in nature, and certainly not over several generations, and will also not provide seed.

The absolute seedless fruit can only be produced by physical, human intervention, other than normal selection. The pollen tubes (of the parent) must be opened by hand, after which the pollen must be removed from the open pollen tubes through scraping, and then, also by hand, applied on the pistil of the tomato plant to be fertilized. In particular, the first two of the abovementioned three steps are unique and totally unnatural.

The invention therefore in a first aspect relates to a method for producing a seedless tomato, a plant carrying seedless tomatoes or capable of carrying seed- less tomatoes, or cultivation material such as seed for such a tomato plant, comprising a. providing a first tomato plant, that is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs, fs); b. providing a second tomato plant, that is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility

(fs,fs); c. crossing the first and second tomato plant for the production of cultivation material, such as seed, which is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs,fs); d. optionally the cultivation of the cultivation material thus obtained into a tomato plant capable of carrying the seedless tomatoes; e. optionally cultivating said tomato plant until it carries the seedless tomatoes, and harvesting the seedless tomatoes thus obtained. More specifically, the invention relates to such a method comprising a. providing a first tomato plant, that is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs,fs) and further is homozygote dominant with respect to at least one property, which is desirable for breeding tomatoes; b. providing a second tomato plant, which is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility

(fs,fs) and is further homozygote recessive with regard to at least one property desired for the breeding of tomatoes; c crossing the first and second tomato plant for the production of cultiva- tion material, such as seed, which is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs,fs), and heterozygote with regard to at least one property desired for the breeding of tomatoes; d. optionally cultivating the cultivation material thus obtained into a tomato plant capable of carrying the seedless tomatoes; e. optionally the cultivating of said tomato plant until it carries the seedless tomatoes, and harvesting the seedless tomatoes thus obtained.

The cultivation material obtained in step c is preferably hybrid seed, so that (also) the tomato plant obtained from said cultivation material in step d is a hybrid tomato plant, with the ensuing advantages known per se associated with the known effect of heterosis. Hereinbelow, under the term "hybrid", both this hybrid cultivation material as well as these hybrid tomato plants are comprised, regardless of whether the latter (already) carry the seedless tomatoes according to the invention. By the combination of the recessive phenotype of parthenocaφy (pk,pk) and the recessive phenotype of functional sterility (fs,fs) these hybrids do not self- pollinate. Nevertheless, the hybrids will form the desired, regularly formed seedless fruits, without the need of (self)-pollination/fertilization of the (pre-embryo's on the) hybrid plant (which according to the invention is neither carried out nor required).

The first and second tomato plant provided in step a and b respectively will hereinbelow be denoted as, and will belong to, the parent lines, and are preferably genetically stable, such as obtained by cultivation/inbreeding over several generations, as further described below. In the above method, the first tomato plant can serve as the "father" (i.e. the plant from which the pollen is obtained) and the second tomato plant can serve as the "mother" (i.e. the plant of which the pistil/the pre-embryo's are fertilized with the pollen of the father), or vice versa. This is not essential for the method of the invention, as in both cases the desired (pk,pk; fs,fs) genotype will be obtained in the hybrid.

The parent lines are preferably chosen such that -besides heterosis- a hybrid with the designed combination of properties is obtained, in a manner analogous to known breeding techniques and cultivation methods for producing hybrids and the desired (combination of) properties therein. These properties are not specifically limited and can comprise all properties which are desired for crossing, cultivating and/or tomato/tomato plants, or for the final tomatoes pro- duced. Examples are early development, increased growth, increased production, the desired form of the plant, the desired form of the fruit, the size of the fruit, the quality of the fruit, etc., as well as increased resistance against diseases, etc. The parent plants are very preferably homozygote with regard to these desired properties (at least one, preferably more), in which one parent plant will be homozygote dominant, and the other plant will be homozygote recessive, and the hybrid will be heterozygote with regard to said property, as is generally known per se for obtaining hybrid tomato plants by breeding.

The number, as well as the nature of the properties, for which the one parent will be homozygote recessive and the other will be homozygote dominant, will preferably be chosen such, that the desired effect of heterosis is obtained in the hybrid. For this puφose, for each of the several properties, the homozygote dominant and homozygote recessive genotypes can be distributed over the parent plants in any desired combination, which means that one parent plant can be homozygote dominant with respect to one desired property, and homozygote recessive with respect to another property, in which case the other parent plant will be homozygote recessive and homozygote dominant respectively, with respect to these properties. All this will be clear to the person skilled in the art of plant breeding. It will also be clear to the skilled person that the uniforπr^'ty of the hybrid obtained according to the invention, and especially of the hybrid seed according to the invention, which is to be marketed, will be essential for both the seed producer as well as the tomato producer, so that a uniform quality of the cultivation material, the hybrid plants and the eventually produced tomatoes can be assured in a reproducible and reliable manner, especially in/for production on a large scale. The uniformity and quality of the seedless tomatoes eventually produced are also very important to the final user, i.e. both the industrial producer of tomato products, as well as the separate consumers.

Such a uniformity of the hybrid can, as known in the art of classical plant breeding, only be obtained in a repeatable and reliable manner, when "pure" parent lines, obtained by inbreeding over several generations, are used. The use of such inbred lines is generally also required for obtaining heterosis, for which the parent lines generally must be genetically "far apart" and (also) after crossing should give a heterozygote genotype for (at least part of) the desired properties in the hybrid produced. The parent lines used must however both be homozygote recessive with regard to both parthenocaφy (pk,pk) as functional sterility (fs,fs), to assure that the hybrid is also reproducibly/reliably provided with the genotype of homozygote recessive parthenocaφy (pk,pk) and homozygote recessive functional sterility (fs,fs), which is essential according to the invention. The presence of the (pk,pk; fs,fs) genotype in the parent lines also means that they are not capable of reproduction by themselves, i.e. without human intervention as described above. This means that said human intervention is not only required for obtaining the hybrids, but also for obtaining and inbreeding the genetically stable parent lines themselves, as well as the (genetically stable) maintenance of the parent lines within the business of the seed producer, so as to enable reproducible and reliable production of the hybrid seed over several generations of the parent lines.

The skilled person, based upon the present description, will be capable of obtaining the desired (combination of) properties as well as the required (pk,pk; fs,fs) genotype in the parent plants. The methods used by the skilled person for this puφose will in general be analogous to known breeding techniques and cultivation methods, with the proviso that for the cultivation over several generations, and in particular the inbreeding of the parent lines, (also) human intervention as described herein will be required (in particular for step c). For cultivating, inbreeding and/or maintaining the parent lines, the pollen obtained from a flower/plant, will however be applied to the same flower or plant, or optionally to a genetically equivalent plant from the same line, in a manner analogous to self- pollination in known inbreeding techniques.

In obtaining the parent lines, it is possible to "cross in" the desired properties from other lines, also in a manner analogous to known breeding techniques. In this way, the desired properties can be crossed into, and obtained in, the parent lines, and thereby in the final hybrid.

The tomato plants obtained in the manner described hereinabove, including the cultivation material such as seeds for these plants, which are suited for use as parent plants in the method described above for obtaining seedless hybrids, as well as the method for obtaining said parent line(s) form further aspects of the present invention.

The crossing of the first and second tomato plant in step c above is carried out by fertilizing the pre-embryo's of the flower of the first or second tomato plant respectively, with pollen obtained from the second or first tomato plant, respectively, dependent upon the choice of the father and the mother.

Because the pollen tube of the father plant is closed, as a result of the functionally sterile phenotype, the pollen must be provided by opening said pollen tube, by machine or preferably by hand, in practice by cutting or scissoring the pollen tube.

Thereafter, the pollen is removed from the pollen tube, preferably again by hand, for instance by scraping, after which the pollen thus obtained is applied to the flower/pistil of the mother plant, again preferably by hand, such as by brushing or another suitable manner, such as spraying, to fertilize the (pre- embryo's of the) mother plant.

Theoretically, it is also possible - after opening the pollen tube, so that the enclosed pollen is released/accessible for spreading/distribution - to use other methods to transfer the pollen onto the pistil of the mother plant, such as using insects, especially when the pollen tube of the mother plants have not been opened or the stamen of the mother plant have been removed to prevent fertilization of the mother plant with its own pollen. Manual fertilization, as described hereinabove, is however very much preferred, both for reasons of efficiency, as well as to avoid fertilization/contamination of the mother. Moreover, manual pollination is a commonly used technique for obtaining hybrid tomatoes, to which for the puφoses of the invention only the opening and scraping of the pollen tubes has to be added. After pollination, the mother plant can be further cultivated until it carries tomatoes, which contain the hybrid seed of the invention. This hybrid seed can then be collected in a manner known per se, optionally be processed further, as well as packaged for storage, transport or sale. Said hybrid seed, optionally in packaged form, forms an important aspect of the invention from a commercial point of view, as will be clear to the skilled person.

For obtaining the hybrid tomato plant and the seedless tomatoes, the hybrid seeds can be sown in a manner known per se, or be made to germinate in another manner known per se in the art, and then be cultivated to tomato plants, which (can) carry the seedless tomatoes of the invention. As mentioned herein- above no (self)-pollination/fertilization of the hybrid occurs; nevertheless a regularly formed fruit is produced on the not-fertilized hybrid.

The seedless tomatoes thus obtained can then be harvested and marketed and/or consumed as such, optionally after one or more further processing steps, such as sorting, washing or packaging.

The invention therefore in further aspects relates to (preferably hybrid) cultivation material for tomatoes such as seed (preferably hybrid) tomato plants, as well as seedless tomatoes obtained and/or obtainable as described hereinabove, and/or suited for use in the method(s) described herein. The seedless tomatoes according to the invention can also be processed further in a manner known per se to tomato products, in particular food products, which may or may not be in a form ready or suited for final use. In this respect, the tomatoes according to the invention have the advantage that they can be processed directly, without a further step for removing the seeds/pips in the pro- duction process.

The invention in a further aspect therefore relates to products, in particular food products, obtained from the seedless tomatoes according to the invention, as well as to a method for obtaining said food products, in which the tomatoes are processed to these products without a separate step for removing the seeds. Such a method can therefore -inter alia- comprise pureeing or mashing in another way of the tomatoes, optionally followed by incoφorating or adding further desired ingredients, and packaging the tomato product thus obtained, without seeds or the residues thereof, in suitable containers for storage, transport or sale, in which said method does not comprise a step for removing any pips/seeds between the mashing of the tomatoes and the packaging of the product. Although the invention has been described hereinabove with respect to a method in which the seedless tomatoes, the tomato plants carrying said seedless tomatoes, and the seeds for producing these plants, are obtained by fertilization of a suited mother plant with pollen of a suited father plant, by means of human intervention, both as described hereinabove, which method is preferred from a practical and economical point of view, it is to be understood that the seedless tomatoes according to the invention, and in particular the plants containing these tomatoes and the seeds for producing these plants, can also be obtained in another suitable manner, such as through recombinant techniques, as well as other suitable methods, such as the asexual reproduction of the tomato plants carrying the seedless tomatoes according to the invention, in a manner known per se.

The method according to the invention can also be used in other plant species or varieties belonging to the family of Solanaceae (night shades).

Claims

C L A I M S

1. Method for obtaining a seedless tomato, a tomato plant carrying said seedless tomatoes or capable of carrying said seedless tomatoes, or cultivation material such as seed for such a tomato plant, comprising a. providing a first tomato plant, that is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs, fs); b. providing a second tomato plant, that is homozygote recessive with regard to the properties parthenocaφy (ρk,pk) and functional sterility (fs,fs); c. crossing the first and second tomato plant for the production of cultivation material, such as seed, which is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs,fs); d. optionally the cultivation of the cultivation material thus obtained into a tomato plant capable of carrying the seedless tomatoes; e. optionally cultivating said tomato plant until it carries the seedless tomatoes, and harvesting the seedless tomatoes thus obtained.

2. Method according to claim 1 , comprising a. providing a first tomato plant, that is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs,fs) and further is homozygote dominant with respect to at least one property, which is desirable for breeding tomatoes; b. providing a second tomato plant, which is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility

(fs,fs) and is further homozygote recessive with regard to at least one property desired for the breeding of tomatoes; c. crossing the first and second tomato plant for the production of cultivation material, such as seed, which is homozygote recessive with regard to the properties parthenocaφy (pk,pk) and functional sterility (fs,fs), and heterozygote with regard to at least one property desired for the breeding of tomatoes; d. optionally cultivating the cultivation material thus obtained into a tomato plant capable of carrying the seedless tomatoes; e. optionally the cultivating of said tomato plant until it carries the seedless tomatoes, and harvesting the seedless tomatoes thus obtained.

3. Method according to claim 1 or 2, in which the seed obtained in step c is hybrid seed and/or in which the tomato plant obtained in step d, which carries or is capable of carryign the seedless tomatoes, is a hybrid tomato plant.

4. Method according to any of claims 1-3, in which step c comprises the following steps: cl . providing pollen derived from the first or second tomato plant, respectively, c2. fertilizing the (pre-embryo's of the) second or first tomato plant respectively, with the pollen obtained in step cl .

5. Method according to claim 4, in which step cl comprises collecting the pollen from the pollen tube of the first or second tomato plant respectively, which is closed due to the presence of the homozygote recessive property of functional sterility (fs.fs).

6. Method according to claim 5, in which the collecting of the pollen in step cl is carried out manually, preferably by cutting or scissoring the pollen tube, followed by scraping the pollen tube.

7. Method according to any of claims 1-6, in which the pollen of the first or second tomato plant respectively, is manually applied to the pistil of the flower of the second or first tomato plant respectively.

8. Method according to any of claims 1-7, in which the tomato plant that carries or is capable of carrying the seedless tomatoes, and that is obtained in step c from the first and second tomato plant, is a hybrid.

9. Cultivation material for tomatoes, such as seed, obtained or obtainable by crossing the first and second tomato plant according to any of claims 1-8, which seed after sowing and culturing provides a tomato plant that carries or is capable of carrying seedless tomatoes.

10. Cultivation material for tomatoes such as seed, in particular according to claim 9, being hybrid cultivation material/seed.

1 1. Tomato plant, obtainable or obtained by crossing the first and second tomato plant according to any of claims 1-8, and/or by sowing and culturing the tomato seed according to claim 9 or 10, which tomato plant carries or is capable of carrying seedless tomatoes.

12. Tomato plant according to claim 11 , being a hybrid.

13. Seedless tomato, obtainable or obtained according to the method of any of claims 1-8, from the tomato seed according to claim 9 or 10, or using a tomato plant according to claim 11 or 12.

14. Processed product, in particular processed food product, on the basis of tomatoes, optionally ready for final use, obtained from the seedless tomatoes according to claim 13.

15. Method for obtaining processed products, in particular food products, on the basis of tomatoes, optionally ready for final use, according to claim 14, comprising providing seedless tomatoes according to claim 13 and processing such to products on the basis of tomatoes in a manner known per se, without a separate step for removing the seeds.

16. Tomato plant, including cultivation material such as seed for such a tomato plant, which is homozygote recessive with respect to the properties of parthenocaφy (pk,pk) and functional sterility (fs,fs) and preferably further homozygote with respect to at least one property desired from the breeding of tomatoes, and which is suited for use as a first or second tomato plant in a method according to any of claims 1-8.

17. Tomato plant according to claim 16, suited for providing hybrid cultivation material according to claim 10 or a hybrid tomato plant according to claim 11.

18. Method for providing a tomato plant according to claim 16 or 17, comprising a. providing a tomato plant that is homozygote recessive with respect to the properties parthenocaφy (pk,pk) and functional sterility (fs,fs) and preferably further homozygote with respect to at least one property desired for the breeding of tomatoes, more preferably a tomato plant according to claim 16 or 17; b. collecting pollen therefrom, preferably by cutting or scissoring the pollen tube, followed by scraping of the pollen tube; c. fertilizing the same or a genetically equivalent tomato plant with the pollen collected in step b; d. collecting the seed from the tomatoes obtained from the fertilized plants of step c; e. optionally cultivating the seed thus obtained to a tomato plant, preferably a tomato plant according to claim 16 or 17; f. optionally repeating steps a-e until a genetically stable parent line is obtained.